Abstract:

Objective　To investigate the effect of anthocyanin (ACN) on improving airway inflammation in asthma model mice by regulating the extracellular matrix metalloproteinase inducer (CD147)/matrix metalloproteinase-9 (MMP-9) pathway. Methods　Male Balb/c mice aged 7-8 weeks, with a weight of (20±5) g, were used to construct the asthma model. The mice were divided into an ovalbumin (OVA) group (model group), L-ACN, M-ACN, and H-ACN groups [75, 150, and 300 mg/(kg·d) of ACN orally administered 1 hour before immune challenge], and an ACN+sCD147 group [300 mg/(kg·d) of ACN orally administered 1 hour before immune challenge + intraperitoneal injection with 22 mg/kg of soluble CD147 (sCD147)], with 10 mice in each group. Another 10 mice were randomly selected from the raised mice as a control group. The airway hyperresponsiveness (AHR) and inflammatory cells in bronchoalveolar lavage fluid (BALF) of the mice in each group were measured. HE staining and PAS staining were applied to observe the pathological changes in lung tissue of asthmatic mice. Enzyme-linked immuno sorbent assay (ELISA) was applied to detect the levels of interleukin (IL)-4, IL-5, IL-13, and interferon (IFN)-γ in the serum of asthmatic mice. Western blot was applied to detect the protein expressions of CD147 and MMP-9 in the lung tissue of asthmatic mice. One-way analysis of variance and SNK-q test were used. Results　In the OVA group, severe cell infiltration was observed in the walls and interstitium of the alveoli, with a significant increase in PAS positive epithelial cells, extensive proliferation of goblet cells, and increased mucus secretion. The AHR, eosinophils (EOS), macrophages (TAM), lymphocytes (Lym), neutrophils (Neu), IL-4, IL-5, IL-13, and protein expressions of CD147 and MMP-9 in the OVA group were higher than those in the control group, and the expression of IFN-γ was lower than that in the control group, with statistically significant differences (all P<0.05). In the L-ACN group, M-ACN group, and H-ACN group, the inflammation of lung tissue was alleviated, the number of PAS positive epithelial cells decreased in sequence, and the mucus secretion decreased. The AHR, EOS, TAM, Lym, Neu, IL-4, IL-5, IL-13, and protein expressions of CD147 and MMP-9 in the L-ACN group, M-ACN group, and H-ACN group were lower than those in the OVA group, and the expression of IFN-γ was higher than that in the OVA group (all P<0.05). In the ACN+sCD147 group, the inflammation of lung tissue was further worsened, the PAS positive epithelial cells increased, and the mucus secretion increased. The AHR, EOS, TAM, Lym, Neu, IL-4, IL-5, IL-13, and protein expressions of CD147 and MMP-9 in the ACN+sCD147 group were higher than those in the H-ACN group, and the expression of IFN-γ was lower than that in the H-ACN group (all P<0.05). Conclusion　ACN can inhibit the CD147/MMP-9 pathway and improve airway inflammation in asthma model mice.

Key words:

Anthocyanin, Extracellular matrix metalloproteinase inducer, Matrix metalloproteinase-9, Asthma, Airway inflammation, Mice, Animal experiment

摘要：

目的　探讨花青素（ACN）调控细胞外基质金属蛋白酶诱导剂（CD147）/基质金属蛋白酶-9（MMP-9）通路改善哮喘模型小鼠气道炎症的作用。方法　采用7～8周龄雄性Balb/c小鼠，体重（20±5）g构建哮喘模型，将其分为卵清蛋白（OVA）组（模型组）、L-ACN组、M-ACN组、H-ACN组［激发免疫前1 h分别灌胃75、150、300 mg/（kg·d） ACN］、ACN+sCD147组［激发免疫前1 h灌胃300 mg/（kg·d） ACN+腹腔注射22 mg/kg可溶性CD147（sCD147）］，每组10只。另从饲养的小鼠中随机选取10只小鼠作为对照组。测定各组小鼠的气道高反应性（AHR）和支气管肺泡灌洗液（BALF）中炎症细胞；HE染色、PAS染色观察哮喘小鼠肺组织病理改变；酶联免疫吸附试验（ELISA）检测哮喘小鼠血清中白细胞介素（IL）-4、IL-5、IL-13和干扰素（IFN）-γ含量；Western blot检测哮喘小鼠肺组织中CD147、MMP-9蛋白的表达。采用单因素方差分析、SNK-q检验。结果　OVA组管壁及肺泡间质出现严重细胞浸润，PAS阳性上皮细胞明显增加，大量杯状细胞增生和黏液分泌增多，AHR、嗜酸性粒细胞（EOS）、巨噬细胞（TAM）、淋巴细胞（Lym）、中性粒细胞（Neu）、IL-4、IL-5、IL-13、CD147、MMP-9蛋白表达均高于对照组，IFN-γ表达低于对照组，差异均有统计学意义（均P<0.05）。L-ACN组、M-ACN组、H-ACN组肺组织炎症减轻，PAS阳性上皮细胞依次减少，黏液分泌减少，AHR、EOS、TAM、Lym、Neu、IL-4、IL-5、IL-13、CD147、MMP-9蛋白表达均低于OVA组，IFN-γ表达均高于OVA组（均P<0.05）。ACN+sCD147组肺组织炎症进一步加重，PAS阳性上皮细胞增加，黏液分泌增加，AHR、EOS、TAM、Lym、Neu、IL-4、IL-5、IL-13、CD147、MMP-9蛋白表达均高于H-ACN组，IFN-γ表达低于H-ACN组（均P<0.05）。结论　ACN可以抑制CD147/MMP-9通路，改善哮喘模型小鼠气道炎症。

关键词:

花青素, 细胞外基质金属蛋白酶诱导剂, 基质金属蛋白酶-9, 哮喘, 气道炎症, 小鼠, 动物实验